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UK Health Security Agency

CHO/dhFr-AC-free

CHO/dhFr-AC-free

Catalogue No.

05011002

Cell Line Name

CHO/dhFr-AC-free

Cell Line Description

This is a derviative of the CHO/dhFr- cell line, ECACC catalogue no. 94060607 which has been adapted to grow in the animal component free medium CHO DHFR (Sigma C8862). The CHO/dhFr- cell line lacks the enzyme dihydrofolate reductase (DHFR) which is necessary for purine synthesis. In the absence of exogenous purines, this enzyme is required for growth. Hence the CHO/dhFr- cell line has growth requirements for hypoxanthine (or adenine), glycine and thymidine. However, the cells will continue to grow without the addition of hypoxanthine and thymidine because the cells can obtain sufficient from the medium. Alpha MEM minus does not contain the ribonucleosides or the deoxyribonucleosides, and will not support the growth of CHO/dhFr- or its serum free adaptation as neither contains the gene to produce DHFR. (See attached data sheet).

Characteristics

Tissue of Origin

Ovary

Karyotype

Not specified

Disease

None Stated

Culture Conditions

Subculture Routine

Maintain cultures between approximately 3-9x10⁵ cells/ml; 5% CO₂; 37°C. Cells grow as aggregates in suspension. Viability may be poor on resuscitation and may initially decrease further. Full recovery may take up to 2 weeks. A centrifugation step to remove the cryoprotectant is essential. Rapidly thaw the frozen ampoule in a water bath at 37°C for 1-2 minutes. Transfer the contents to a centrifuge tube and slowly add 5-10ml of pre-warmed growth media. Remove a sample for counting. Centrifuge at 100g for 2-3 minutes to pellet cells and seed at a relatively high density of 5-7 x 10⁵ cells/ml. Leave culture flask upright and observe regularly until viable proliferating cells are seen. Once established use a split ratio of 1:2 approximately every 4 to 5 days.

Culture Medium

CHO DHR medium (Sigma C8862) + 2mM Glutamine + 100uM Hypoxanthine + 16uM Thymidine (e.g. use HT supplement Sigma H0137). Addition of Aminopterin (Sigma A5159) 0.4uM is also advised. When freezing cells use freeze medium composed of 50:50 fresh:conditioned medium + 10% DMSO.

Growth Mode

Aggregates in suspension

Additional Info

Depositor

Dr. E. Penn, European Collection of Cell Cultures, Health Protection Agency, Porton Down, Salisbury, SP4 0JG, UK

Country of Origin

United Kingdom

Hazard Group (ACDP)

Hazard Group (ACDP) 2

Applications

References

None Specified By Depositor

Available Formats

  • Frozen
  • DNA-5µg (100ng/µl)

If use of this culture results in a scientific publication, it should be cited in the publication as: CHO/dhFr-AC-free (ECACC 05011002).

Unless specified otherwise, at ECACC we routinely handle all of our cell lines at containment level 2 in accordance with the ACDP guidelines (Advisory Committee on Dangerous Pathogens) (UK). All cell cultures have the potential to carry as yet unidentified adventitious agents. It is the responsibility of the end user to ensure that their facilities comply with biosafety regulations for their own country. ACDP Guidance: Biological agents: Managing the risks in laboratories and healthcare premises.

The Culture Collections represent deposits of cultures from world-wide sources. While every effort is made to ensure details distributed by Culture Collections are accurate, Culture Collections cannot be held responsible for any inaccuracies in the data supplied. References where quoted are mainly attributed to the establishment of the cell culture and not for any specific property of the cell line, therefore further references should be obtained regarding cell culture characteristics. Passage numbers where given act only as a guide and Culture Collections does not guarantee the passage number stated will be the passage number received by the customer.

Cultures supplied by Culture Collections are for research purposes only. Enquiries regarding the commercial use of a cell line are referred to the depositor of the cell line. Some cell lines have additional special release conditions such as the requirement for a material transfer agreement to be completed by the potential recipient prior to the supply of the cell line. Please view the Terms & Conditions of Supply for more information.